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. Author manuscript; available in PMC: 2009 Oct 6.
Published in final edited form as: Eur J Immunol. 2008 Oct;38(10):2697–2705. doi: 10.1002/eji.200838186

Plasmodium vivax parasites alter the balance of myeloid and plasmacytoid dendritic cells and induction of regulatory T cells

Kulachart Jangpatarapongsa 1,2,*, Patchanee Chootong 1,2,*, Jetsumon Sattabongkot 3, Kesinee Chotivanich 4, Jeeraphat Sirichaisinthop 5, Sumalee Tungpradabkul 6, Hajime Hisaeda 7, Marita Troye-Blomberg 8, Liwang Cui 9, Rachanee Udomsangpetch 1,2
PMCID: PMC2757553  NIHMSID: NIHMS138525  PMID: 18825754

Summary

Immunity induced by Plasmodium vivax infections leads to memory T cell recruitment and activation during subsequent infections. Here, we investigated the role of regulator T cells (Treg) in coordination with the host immune response during P. vivax infection. Our results showed a significant increase in the percentage of FOXP3+ Treg, IL-10 secreting Type I Treg (Tr1) and IL-10 levels in patients with acute P. vivax infection as compared to those found in either naïve or immune controls. The concurrent increase in the Treg population could also be reproduced in vitro using PBMC from naïve controls stimulated with crude antigens extracted from P. vivax-infected red blood cells. Acute P. vivax infections were associated with a significant decrease in the numbers of DC, indicating a general immunosuppression during P.vivax infections. However, unlike P. falciparum infections, we found that the ratio between myeloid DC (MDC) and plamacytoid (PDC) was significantly lower in acute P. vivax patients than that of naïve and immune controls. Moreover, the reduction of PDC may be partly responsible for the poor antibody responses during P. vivax infections. Taken together, these results suggest that P. vivax parasites interact with DC, which alters MDC/PDC ratio that potentially leads to Treg activation and IL-10 release.

Keywords: Plasmodium vivax, malaria, immunity to malaria, regulatory T cell, IL-10, dendritic cell

1. Introduction

Malaria is a common tropical disease causing deaths among Plasmodium falciparum-infected children mainly in Sub-Saharan Africa [1]. P. falciparum causes malignant tertian malaria that accounts for most malaria-associated deaths, whereas P. vivax causes relapsing fever and the infection rarely becomes fatal. Although a better understanding of immunity is needed for the design of effective vaccines, immune regulation in the host during malaria infection is not fully understood, and few studies have been conducted in patients with P. vivax infections. Our recent study has shown that anti-P. vivax antibody levels were very low in immune individuals living in endemic area and in patients with acute P. vivax malaria. For the cell-mediated arm, an acute P. vivax infection was associated with the activation of memory T cells belonging to either a cytotoxic or helper phenotype [2]. Additionally, previous evidence [3, 4] shows that immunization with pre-erythocytic antigens can induce IFN-γ release. This suggests that P. vivax can activate the immune system via the Th1 pathway. However, a possible suppressing mechanism arise from the activation of regulatory T cells (Treg) as has been shown in a murine malaria study [5].

Treg constitutively express CD25, which is the IL-2/α chain receptor [6]. Co-presentation of CD25 with fork head box protein P3 (FOXP3) dictates the immune-suppressive role of Treg via the release of IL-10 and TGF-β [7]. Treg have been shown to alter the balance between myeloid dendritic cells (MDC) and plasmacytoid dendritic cells (PDC) in blood, which eventually determines the outcome of T cell responses [8]. The increase in MDC/PDC ratio is associated with the activation of the Th1 pathway, whereas a decreased ratio is associated with the activation of the Th2 pathway [8–10]. In a murine malaria model, the parasites have been shown to evade the immune response via activation of Treg [11]. A recent study confirms this trend during P. falciparum infections where Treg activation was correlated with higher rates of parasite growth [12]. However, the role of Treg during the course of P. vivax infection has not been investigated. Here, we compared Treg population and IL-10 levels among acute P. vivax patients, immune and naïve controls. Results revealed significant association between of vivax infections with increased levels of Treg and IL-10. In contrast, P. vivax infections led to a general reduction in dendritic cells (DC) and lowered MDC/PDC ratio, suggesting a possible immunosuppressive role of Treg during acute P. vivax infections.

2. Results

2.1 Activation of Treg during P. vivax infection

The percentage of CD4+CD25+ T cells was quantified and the results are shown in Fig. 1A. Immune individuals living in endemic areas had a similar level of Treg as observed in naïve controls. However, the level was significantly elevated during acute P. vivax infections (mean, quartile percentages=13.8%, 11.3–16.2) when compared to that of naive controls (7.5%, 6.3–8.8) [P=0.001], and immune controls (4.3, 4.7–9.9) [P=0.001].

Figure 1.

Figure 1

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Increased numbers of FOXP3+ Treg in patients with acute P. vivax infections. Flow cytometric analyses of cryopreserved PBMC stained with anti-CD4 Tricolor, anti-CD25 PE, and anti-FOXP3 Alexa Fluor® 488 from the indicated groups and lymphocyte populations were gated. (A) CD4+CD25+ T cells in the lymphocyte population are indicated by the boxed areas. (B) Histograms of FOXP3 expression in the CD4+CD25+ T cells. The percentages of CD4+CD25+ (A) and CD4+CD25+FOXP3+ (B) cells are shown. All figures show one representative sample out of seventeen.

The expression of FOXP3+ on CD4+CD25+ T cells was analyzed to differentiate the FOXP3+ Treg [13]. The results showed that FOXP3+ Treg in the acute P. vivax malaria patients (20.8%, 12.9–68.3) were approximately 7- and 3-fold higher than that of immune controls (2.7%, 1.9–4.6) [P<0.001] and naïve controls (6.3%, 2.4–10.2) [P<0.001], respectively. Interestingly, the level of FOXP3+ Treg in the malaria immune controls were significantly lower than that seen in naive controls [P=0.4] (Fig. 1B).

2.2 IL-10 and the activation of Treg

It is known that Treg produce the anti-inflammatory cytokine IL-10. Results indicated that IL-10 was 4-fold higher during acute P. vivax infections than those measured in immune controls [P<0.001], and 2-fold higher than that of naïve controls [P=0.005] (Fig. 2A). The levels of IL-10 in plasma collected from immune controls were significantly lower than that seen in naive controls [P=0.008]. As expected, the level of IL-10 producing Treg (CD4+CD25+IL-10+ or type 1 Treg or “Tr1” cells) of P. vivax infected patients were also significantly higher than that of immune controls [P=0.001] and naïve controls [P=0.004] (Fig. 2B). Moreover, there was a significant association in the numbers of FOXP3+ Treg and Tr1 cells in acute P. vivax patients (R2=0.3, P=0.006) and immune controls (R2=0.4, P=0.01). This association was not found in naïve controls (Fig. 2C).

Figure 2.

Figure 2

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Comparison of the levels of plasma IL-10 (A) and Tr1 cells (B) between naive controls (NC), immune controls (IC) and patients acutely infected with P. vivax (AC). The correlation between Tr1 cells and FOXP3+ Treg in lymphocytes from patients acutely infected with P. vivax (C). Data are shown as mean (thick horizontal line), inter-quartile range (box plot), maximum and minimum (upper-lower lines) (A, B). The numbers represent mean, inter-quartile range. (Data were log transformed and independent sample t test was used to calculate P value)

2.3 Regulatory-T cell stimulation by P. vivax antigens

Infection with P. vivax resulted in an increase levels of Treg and plasma IL-10. To reproduce these phenomena in vitro, PBMC from naïve controls were stimulated with P. vivax antigens and the numbers of FOXP3+ Treg were determined on day 5 (Fig. 3A). The results indicated that the number of FOXP3+ Treg were significantly increased when PBMC was stimulated with 50 µg/ml of P. vivax antigen as compared with that of Normal RBCs (NRBC) [P=0.01] and that of PHA [P=0.01]. Consistent with ELISA results, the levels of Tr1 cells by as measured from flow cytometric analysis was found to increase after stimulation with 50 µg/ml of P. vivax antigen as compared with that of NRBC [P=0.048] (Fig. 3B). Moreover, RT-PCR we also detected the expression of IL-10 in naïve PBMC after 5 days of stimulation with P. vivax antigens (data not shown).

Figure 3.

Figure 3

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Increased numbers of FOXP3+ Treg (A) and Tr1 cells (B) in lymphocytes after stimulation with P. vivax antigens. PBMC from naïve controls cultured with Normal red blood cells or PV schizont lysate at the indicated concentrations were analyzed for flow cytometric analysis. Data represent mean percentage ± SD of five experiments. Data were log transformed and independent sample t test was used to calculate P value.

2.4. Blood dendritic cells during acute P. vivax infection

Circulating DC defined as HLA-DR+MDC and HLA-DR+PDC were determined in acute P. vivax patients and naïve controls (Fig. 4A–C). The results showed that the absolute numbers of both MDC and PDC were significantly reduced in P. vivax patients than those of naïve controls [P =0.09, and 0.03, respectively] (Fig. 4D). Consequently, the MDC/PDC ratio was significantly different between naïve controls and acute P. vivax patients [P=0.01] (Fig. 4E).

Figure 4.

Figure 4

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(A–C) Flow cytometric analyses of whole blood stained with anti-HLA-DR PerCP and Lin FITC, anti-CD11c RPE (for MDC), or anti-CD123 RPE (for PDC). (R2 showed the population of HLA-DR+Lin−, I inserted an arrow in the Figure 4A) (D) Number of MDC and PDC, (E) MDC/PDC ratio in the peripheral blood of patients acutely infected with P. vivax (AC) and naive controls (NC). Numbers represent mean ± SD. (n=?? Acute infections=17 and naïve controls=15) Sample t test was used to calculate P value.

During infection, P. vivax parasites may activate Treg via P. vivax-primed MDC or PDC of the host. Therefore, the association between MDC or PDC with Treg was analyzed. However, there was no association between PDC or MDC with FOXP3+ Treg or Tr1 cells (data not shown).

3. Discussion

Infections by pathogens can lead to either the Th1 or Th2 response. Th1 response often leads to the resolution of malaria infection, and therefore, is favored in vaccine design [14]. IFN-γ produced by CD8 T cells and/or NK cells inhibits parasite development, thereby contributing to the protection against pre-erythocytic stages of both P. falciparum and P. vivax [15]. However, there are studies showing the induction of IL-10 in malaria patients, suggesting that Th2 pathway may also be involved in malaria immunity [16]. Quite different from the findings in P. falciparum, we found that P. vivax infections rarely resulted in significant production of parasite-specific antibodies, but that acute P. vivax infections induced Treg activation. This result is consistent with two recent findings in both P. yoelii and P. falciparum, suggesting that parasites may use a similar mechanism of immune evasion [11, 12]. Although the interactions among malaria-specific T cells, B cells and Treg remain poorly characterized, our result suggested that the increased frequency of Treg during acute P. vivax infection may lead to suppression of both cell- and antibody-mediated immunity, which may benefit parasite survival in hosts. There are evidence indicates that Treg can suppress T-cell responses through the production of IL-10 and TGF-β [17, 18], whereas they may suppress B-cell maturation and differentiation directly [19, 20] or indirectly through the down-regulation of IL-2 or IL-4 production in responder lymphocytes. This may account for low parasite-specific antibody levels seen in P. vivax patients [2].

In this study, patients with acute P. vivax infection were observed to exhibit higher plasma levels of IL-10 than those of naïve and immune controls. Similar findings have been reported in both P. vivax and P. falciparum infections [21–24]. Even though there is evidence that CD8 Treg produce IL-10 to suppress T cell responses [25], our result showed a significant correlation between elevated levels of Tr1 and FOXP3+ Treg, a result further corroborating our earlier finding of increased levels of total CD4+ T cells during acute P. vivax infections [2]. In parallel, we also found a concurrent increase levels of Treg and expression of IL-10 when PBMC from naive controls were stimulated with P. vivax antigens in vitro. This result is also consistent with findings that the number of Treg is increased in cord blood mononuclear cells of placental P. falciparum infections and IL-10 elevation after stimulation with P. falciparum antigens [26]. Altogether, these results suggest that Treg may be a direct source of IL-10 production and/or enhance the IL-10 production by DC [27]. Recent report showing that the stimulated FOXP3+ Treg produces many cytokines including IL-10, resulting in activation of monocytes/macrophages can support the former hypothesis [28]. Our finding showed significant association between different Treg populations during acute P. vivax infection, despite the fact some acutely infected cases (23%) appeared to have elevated proportion of Tr1 cells with comparatively low levels of FOXP3+ Treg. Previous studies have demonstrated that Tr1 cells generated in vitro do not express FOXP3+, but it was found to maintain the suppressor function [29, 30]. This suggests that FOXP3 expression is not a prerequisite for the suppressive function of Tr1 cells. On the other hand, Tr1 cells specific for desmoglein3, the dominant autoantigen in pemphigus vulgaris, are shown to constitutively express FOXP3 [31]. Therefore, the association of Tr1 cells with FOXP3+ Treg remains controversial and needs further investigation [32].

Among the three groups of volunteers, the lowest levels of IL-10 and Tr1 cells were observed in the immune controls. This suggests that activation of Treg is transient during P. vivax infections, and that subsequent decline of Treg after parasite clearance could be due to higher levels of memory T cells that were maintained after the infection [2]. Our study did not find a correlation of CD4+CD25+ and FOXP3 levels. This result is in line with a recent study showing no correlation between CD25+ cells and FOXP3 level in human CD4+ cells [33].

The immunoregulatory effect of malaria infections is related with the down regulation of antigen presenting DC. Consistent with earlier studies documenting the immunosuppressive effect of malaria parasites on the maturation and differentiation of DC [34, 35], we found significantly lower levels of DC (both PDC and MDC) in acute P. vivax patients. This finding coincides with the recent study showing the decreasing numbers of MDC and PDC among children with severe P. falciparum infection [36]. Another reason could be the reallocation of cells away from the peripheral blood, e.g., to the spleen. The evidence of lymphopenia in both P. falciparum and P. vivax infection supports such an explanation [37–39]. The levels of MDC and PDC in the peripheral blood are recovered during post-treatment [36].

It has been shown that the depletion of PDC abrogated the secretion of specific polyclonal IgG in response to influenza virus [40]. Therefore, the reduction of PDC level during P. vivax infection may account for the low level of P. vivax-specific antibodies in vivax patients [2]. This suggests that PDC are critical for the generation of plasma cells and antibody responses during parasite/pathogen infections. However, unlike that of P. falciparum infected patients [36], the percentage of PDC was higher than that of MDC during acute P. vivax infections, resulting in a lower MDC/PDC ratio. This may suggest that P. vivax parasites suppress host immune response through elevated levels of PDC and Treg. Two pieces of evidence show that stimulation of PDC induces naïve CD4 and CD8 T-cell differentiation into Th2 cells and IL-10 producing CD8 T cells [8, 41].

Taken together, we found that acute P. vivax infections were associated with higher levels of Treg and IL-10, but lower levels of DC in the blood. This suggests that Treg are activated by parasites during acute infections, which then play an immunosuppressive role of host cell- and antibody-mediated immunity, resulting in retarded parasite clearance. While the overall levels of DC were reduced, the balance between the two cell types (MDC and PDC) was altered. In addition, our previous study also revealed an increase in the levels of γδ T cells during acute P. vivax infection. Therefore, the interactions among different DC, T cell phenotypes and P. vivax antigens await further investigations.

4. Materials and Methods

4.1 Study population

Blood samples were collected from 17 patients with acute P. vivax infections at two malaria clinics in Mae Sot and Mae Kasa, Tak province, Thailand. Diagnosis of P. vivax malaria infection was based on the examination of Giemsa-stained thick blood films. Polymerase chain reaction (PCR) with four human malaria (P. falciparum, P. vivax, P. malariae, and P. ovale) species-specific primers was performed on DNA isolated from blood samples to verify malaria infections. Only single P. vivax infection was recruited in the experiments. Blood samples were collected from additional 25 people residing in the same P. vivax-endemic area. The subjects defined as “immune controls” have had recent malaria infections and had anti-P. vivax antibodies determined by ELISA. These immune controls did not have P. vivax infections at the time of blood collection as determined by both microscopic and PCR analysis. Fifteen healthy adults living in Bangkok without previous malaria exposure or antibodies to malaria parasites were recruited to serve as “naïve controls”. The clinical characteristics of the subjects are listed in Table 1. This study was approved by the Committee on Human Rights Related to Human Experimentation, Mahidol University, and the Ministry of Health, Thailand. Informed consent was obtained from each individual before the blood sample was taken.

Table 1.

Information and Clinical data of P.vivax patients, immune and naïve controls *

No. of
cases		 Age
(Years)		 Sex
 Hematocrit
(%)		 Temperature
(°C)		 Parasitaemia
(%)
M F
AC 17 38 ± 11 7 10 43 ± 7 37.5 ± 1.3 0.4 ± 0.3
(15 – 52) (35 – 63) (35 – 40) (0.03 – 1.2)
IC 25 38 ± 13 16 9 46 ± 3 36.8 ± 0.4 0
(18 – 77) (43 – 48) (36 – 37)
NC 15 29 ± 7 8 7 42 ± 4 37.0 ± 0.5 0
(22 – 43) (37 – 45)
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*

Mean ± Standard deviation (range).

AC= Acute P. vivax infection, IC=Immune controls and NC=Naïve controls.

4.2 Preparation and cryopreservation of peripheral blood mononuclear cells (PBMC)

Venous blood from P. vivax patients, immune controls, and naïve controls was collected in heparinized tubes and PBMC were separated by gradient centrifugation using Lymphoprep™ (AXIS-Shied PoC AS, Oslo, Norway) according to the manufacturer’s recommendations. The PBMC pellet was resuspended at a concentration of 106 cells/ml in RPMI-1640 supplemented with 10% heat-inactivated fetal bovine serum (FBS) (Invitrogen, Carlsbad, USA). The viability of PBMC, normally above 98%, was determined by tryphan blue exclusion. PBMC in 10% dimethyl sulfoxide (DMSO) were cryopreserved in liquid nitrogen until analysis.

4.3 Parasite cultures and antigen preparations

Crude P. vivax antigens were obtained from P. vivax-infected red blood cells (iRBCs). Briefly, P. vivax infected blood was depleted of white blood cells by filtering through a sterile column of CF11 cellulose (Whatman, Maidstone, UK) and the RBCs were washed with RPMI-1640 by centrifugation at 1190 g for 5 min. The parasites were cultured in an atmosphere of 5% CO2, 5% O2 and 90% N2 for 24 – 30 h at 5% hematocrit in McCoy’s 5A medium (Invitrogen) supplemented with 25% human AB serum until they matured to the schizont stage (≥6 nuclei). These mature parasites were enriched by centrifugation on 60% Percoll (Pharmacia, Uppsala, Sweden) at 1190 g for 10 min. The enriched iRBC pellets were sonicated for 40 sec at 150 watts and the protein concentration was determined by the Bradford assay (Bio-Rad, Hercules, USA). The vials were then aliquoted and stored at −70°C until use. Uninfected RBCs were processed similarly and stored at −70°C to be used as a negative control.

4.4 In vitro stimulation of PBMC

PBMC from five healthy donors were used for in vitro stimulation. PBMC (2 × 105 cells/well) in RPMI-1640 supplemented with 25 mM HEPES, 1.8 mg/ml D-glucose, 2 mM glutamine, 40 mg/ml of gentamicin and 10% heat-inactivated FBS were cultured for 5 days at 37°C in a humidified chamber with 5% CO2, 5% O2 and 90% N2 in the presence of P. vivax antigens at a concentration of 1, 10, or 50 µg/ml. Medium alone, equivalent concentration of RBC extracts or 2 µg/ml of phytohemagglutinine (PHA) were used as negative and positive controls, respectively. All experiments were performed in duplicates. After 5 days of activation, the cells were harvested and stained for Treg marker, CD25 and FOXP3 or IL-10.

4.5 Intracellular staining and flow cytometric (FCM) analysis

For the three-color FCM analysis, PBMC (105 cells/vial) were stained with a combination of fluorochrome-conjugated monoclonal antibodies (mAbs): RPE-Cy5-labeled anti-CD4 (Caltag, Burlingame, USA) and RPE-labeled anti-CD25 (Immunotech, Marseille, France) for 30 min at 4°C and washed with phosphate-buffered saline (PBS). After staining, the cell pellets were fixed with a fixative solution and washed with a permeabilizing solution according to the manufacturer’s recommendations (BioLegend, San Diego, USA). After incubation in the permeabilizing buffer for 20 min at room temperature, the cells were incubated with Alexa fluor® 488-labeled anti-FOXP3 (BioLegend) or FITC-labeled anti-IL-10 for 30 min and then washed with PBS. Cells were fixed with the fixative solution for data acquisition and analysis on FACSCalibur using the CELLQUEST software (Becton Dickinson, San Jose, USA).

4.6 Phenotyping of circulating blood DCs

Two hundred µl of whole blood from P. vivax-infected patients were stained with an antibody mixture containing lineage-specific mAbs to CD3, CD14, CD19, CD20, CD56, and CD66b conjugated with FITC (lin-FITC) (Caltag), antibodies to CD11c (Serotec, Oxford, UK) and CD123 conjugated with RPE (BioLegend), and antibodies to HLA-DR conjugated with PerCP (Becton Dickinson). Stained cells were treated with RBC lysis solution. At least 2 × 105 cells were analyzed in a FACSCalibur flow cytometer (Becton Dickinson). HLA-DR+ CD123+lin− cells were defined as PDC and HLA-DR+CD11c+lin− as MDC. The absolute number of circulating MDC and PDC was obtained by multiplying the percentage of MDC or PDC by the number of leukocytes per milliliter of blood.

4.7 Determination of IL-10 by enzyme-linked immunosorbent assay (ELISA)

Polystyrene immunoplates (Corning, Corning, USA) were coated with 25 µl of 1 µg/ml of anti-human IL-10 mAb (Mabtech, Nacka, Sweden) diluted in PBS (pH 7.4) and incubated overnight at 4°C. Each well was blocked with 50 µl of 0.1% bovine serum albumin for 90 min at room temperature. After five washings with PBS, 25 µl of plasma (1:2 dilution) or PBS control were added into duplicate wells and incubated overnight at 4°C. The plates were then washed for five times with PBS, after which 25 µl of anti-human IL-10 conjugated with biotin (dilution 1:2000) were added and incubated for 90 min at 37°C, followed by the addition of 25 µl of streptavidin-alp-PQ (dilution 1:2000) (Mabtech) at 37°C for 90 min. After five final washings, 25 µl of p-nitro phenylphosphate (pNPP) (Sigma, Saint Louis, USA) were added and incubated for 30–60 minutes in the dark at room temperature. Enzyme activity was measured by an automated microplate reader, V-Max (Molecular Device, Sunnyvale, USA) at 405 nm.

4.8 Determination of IL-10 gene expression by Reverse Transcriptase-PCR (RT-PCR)

RNA was extracted from six samples after in vitro stimulation using Trizol® solution (Invitrogen) according to the manufacturer’s recommendation. Total RNA was reverse transcribed to cDNA, primers of the IL-10 gene and amplification conditions were as described previously [42]. The expression of IL-10 gene (138 bp) was determined in 1.5% agarose gels containing ethidium bromide. Primers of β-actin were designed as described previously [43] and used as a control (790 bp).

4.9 Data analysis

All data were analyzed using the SPSS program (Version 10.0, Chicago, USA). The percentages of CD4+CD25+, FOXP3+ Treg, Tr1 phenotypes, plasma IL-10 levels and number of MDC and PDC, both from volunteers and in vitro stimulation were log transformed to produce normal distributions. Parametric analysis was performed using transformed data as follows: the mean percentage difference for Treg phenotypes, IL-10 levels and number of MDC and PDC between the groups (i.e. naïve controls vs. immune controls, naïve controls vs. acute infection, and immune controls vs. acute infection) were analyzed using the independent samples t test. The Spearman approach was used to evaluate correlation of Tr1 with FOXP3+ Treg, and correlation of MDC or PDC levels with Treg. The results were considered statistically significant (P<0.05) at the 95% confidence interval.

Acknowledgements

We thank all staff at the Mae Sot and Mae Kasa Malaria Clinics, the Department of Entomology, AFRIMS, Bangkok, and the Malaria Training Center, Saraburi, Thailand for collection of the samples. We also thank M. Hagstedt, W. Jangiam for technical support. KJ was a research fellow supported by the Fogarty International Center (FIC), National Institutes of Health (NIH). This work was partly supported by The Thailand Research Fund to RU, by The Commission on Higher Education to KJ (CHE-RES-PD), by BioMalPar to MT and by a grant (D43-TW006571) to LC from FIC, NIH.

Abbreviations

FOXP3

forkhead box protein P3

Treg

Regulatory T cells

FOXP3+ Treg

CD4+CD25+FOXP3+ T cells

Tr1

T Regulatory 1 cells or CD4+CD25+IL-10+ T cells

Footnotes

Conflict of interest: No

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